This protocol aims to evaluate mechanisms governing interrelationships among malaria, intravascular hemolysis, nitric oxide bioavailability, endothelial function, pulmonary hypertension, and evolutionarily-selected host polymorphisms that regulate the host response to hemolysis. We are correlating our clinical observations in the field with laboratory assays of hemolysis and nitric oxide bioavailability related to scavenging by cell-free hemoglobin and arginine catabolism. Using a candidate gene approach, we will identify and selectively characterize polymorphisms in genes important for endothelial function, vascular inflammation and disease phenotype. Finally, the characterization of this mechanism in malaria may catalyze the development of novel therapies targeting this pathway, such as sodium nitrite, inhaled nitric oxide gas, and/or recombinant haptoglobin infusions. This study took place in a malaria-endemic region of Mali in West Africa at Hospital Gabriel Tour in Bamako, which is the major pediatric referral hospital in Mali. The first subject was enrolled on 10/20/08. As of 6/25/08, 102 participants (50 males, 52 females) in Mali had been enrolled into the study. The mean age was 32 months. A standardized history and physical examination was performed. Each participant also underwent echocardiography (165 completed echocardiograms including follow-ups). Obtaining a blood sample for standard hematology and chemistry panels, research bloods, as well as DNA collection for isolation and plasma storage followed the echocardiogram. Based on these studies the prevalence of PAH in the severe malarial anemia subjects was 64% versus 19% in the healthy controls. Children with severe malaria had higher plasma levels of hemoglobin and arginase-1, reduced wholeblood levels of nitrite, and increased NO consumption relative to controls. They also had increased pulmonary arterial pressures (P<.05) with elevated levels of NT-proBNP and soluble vascular cell adhesion molecule-1 (P<.001). Children with severe malaria have increased pulmonary pressures and myocardial wall stress. These complications are consistent with NO depletion from intravascular hemolysis, and they indicate that the pathophysiologic cascade from intravascular hemolysis to NO depletion and its cardiopulmonary effects is activated in children with severe malaria. We also have echocardiographs on the asymptomatic parasitemia and uncomplicated malaria arms, but this data has not yet been analyzed. This year we have performed HPLC analysis of 181 plasma samples from 97 subjects. In summary, we have found arginine depletion and relative elevation of ADMA in plasma of children with acute malaria infection. This confirms our prior observations in West African children. The correlations with nitric oxide production and hemolysis are currently undergoing analysis. Currently we are performing HPLC analysis that includes a full amino acid profile to examine plasma concentration of citrulline and ornithine. Citrulline and Ornithine are major products of two key reactions, the conversion of arginine to citrulline by nitric oxide synthase and the conversion of arginine to ornithine by arginase. Arginase activity is elevated in blood of malaria patients, possibly due to decompartmentalization of arginase from red blood cells, or by the activity of parasite-synthesized arginase. To assess the activity of arginase we wish to measure plasma arginine:ornithine ratio. We will measure the amino acid profile using the Waters MassTrak reagent and a Waters UPLC system.